DESCRIPTION (supplied by applicant) Thyroid hormones (THs) and neuronal excitability are both essential for fetal neurodevelopment. In vitro experiments suggest THs may regulate excitability during embryonic and neonatal stages. This proposal seeks to characterize in vivo TH signaling in embryonic neurons, focusing on voltage-gated sodium current (INa) in zebrafish (Danio rerio) Rohon-Beard sensory neurons (RBs). Preliminary data show application of one type of TH, thyroxine (T4), rapidly increases RB INa amplitude. The rapid time course is not consistent with conventional TH mechanisms involving nuclear receptors. Additionally, the integrin antagonist, echistatin, inhibits the rapid action of T4, suggesting involvement of RGD-type integrin receptors. This proposal aims to outline the novel T4 signaling mechanism in embryonic zebrafish neurons with two specific aims. Aim 1 will determine in vivo relevance of T4 signaling. Preliminary data show blockade of T4 receptors reduces RB INa amplitude relative to controls, suggesting endogenous T4 regulates INa amplitude. To test whether reduction of endogenous T4 affects INa, I will deplete endogenous T4 stores to different extents by yolkectomy. Then I will test whether effects of yolkectomy are specific to T4 by supplementing T4 and/or T3 to yolkectomized embryos. To control for nonspecific effects of yolkectomy I will measure membrane properties not regulated by T4 and voltage-gated potassium current, which preliminary data suggest is not regulated by T4. Aim 2 will identify the specific integrin receptor(s) mediating acute T4 action on RBs. Preliminary data indicate blockade of all RGD type integrin receptors occludes T4's acute action. To determine the specific integrin receptor that mediates T4's acute action I will first determine which integrin subunits are expressed on or near RBs. Second, I will selectively block candidate integrin receptors with function blocking antibodies while recording INa during acute T4 application and without T4 application to determine whether candidate integrins regulate INa in vivo. I will also test whether relevant integrins are located directly on RBs by physically isolating RBs with the nucleated patch method. Overall, this proposal will identify the in vivo role for T4, the identities of specific T4 receptors in the embryonic nervous system. Relevance: The prevalence of maternal hypothyroidism during pregnancy is 3% in the United States. Recent evidence shows maternal hypothyroidism can delay nervous system development in offspring. Understanding how maternal thyroid hormones affect embryonic neurons will help determine how to manage and treat hypothyroid pregnancies.